* U.S. Environmental Protection Agency, National Health and Environmental Effects Research Laboratory, Mid-Continent Ecology Division, Duluth, Minnesota 55804;
Toxicol Sci. 2013 Dec;136(2):359-72. doi: 10.1093/toxsci/kft219. Epub 2013 Oct 4.
Isolated perfused trout livers were used to evaluate in vitro-in vivo metabolism extrapolation procedures for fish. In vitro depletion rates for 6 polycyclic aromatic hydrocarbons (PAHs) were measured using liver S9 fractions and extrapolated to the intact tissue. Predicted hepatic clearance (CLH) values were then compared with values exhibited by intact livers. Binding in liver perfusates was manipulated using bovine serum albumin (BSA) and was characterized by solid-phase microextraction. Additional studies were conducted to develop binding terms (f U; calculated as the ratio of unbound fractions in liver perfusate [f U,PERF] and the S9 system [f U,S9]) used as inputs to a well-stirred liver model. Hepatic clearance values for pyrene and benzo[a]pyrene, predicted by extrapolating in vitro data to the intact tissue, were in good agreement with measured values (< 2-fold difference). This can be partly attributed to the rapid rate at which both compounds were metabolized by S9 fractions, resulting in perfusion-limited clearance. Predicted levels of CLH for the other PAHs underestimated observed values although these differences were generally small (< 3-fold, except for naphthalene). Setting f U = 1.0 improved clearance predictions at the highest tested BSA concentration (10mg/ml), suggesting that trout S9 fractions exhibit lower levels of intrinsic activity than the intact tissue or that the full binding assumption (ie, f U = f U,PERF/f U,S9) underestimates the availability of hydrophobic substrates to hepatic metabolizing enzymes. These findings provide qualified support for procedures currently being used to predict metabolism impacts on chemical accumulation by fish based on measured rates of in vitro activity.
采用离体灌流鳟鱼肝脏，评估多环芳烃（PAHs）在鱼类体内外代谢的外推程序。应用肝脏 S9 部分测量 6 种多环芳烃的体外耗竭率，并将其外推至完整组织。然后将预测的肝清除率（CLH）值与完整肝脏的表现值进行比较。通过牛血清白蛋白（BSA）操纵肝灌流中的结合，并通过固相微萃取进行特征描述。还进行了进一步的研究，以开发用于输入搅拌良好的肝模型的结合项（fU；计算为肝灌流中未结合分数[fU，PERF]与 S9 系统[fU，S9]的比率）。通过将体外数据外推至完整组织来预测的芘和苯并[a]芘的肝清除率值与测量值吻合较好（<2 倍差异）。这部分归因于 S9 部分快速代谢这两种化合物，导致灌流限制清除率。尽管这些差异通常较小（<3 倍，萘除外），但预测的其他 PAHs 的 CLH 值低估了观察值。在最高测试的 BSA 浓度（10mg/ml）下，将 fU 设置为 1.0 可改善清除率预测，这表明鳜鱼 S9 部分表现出比完整组织更低的固有活性，或者完全结合假设（即 fU=fU，PERF/fU，S9）低估了疏水性底物对肝代谢酶的可用性。这些发现为目前基于体外活性测量值预测鱼类对化学物质积累的代谢影响的程序提供了有条件的支持。
